The long-term objective of this proposal is to develop novel therapeutics for periodontitis by targeting cathepsin K (Ctsk) and attenuating Toll-like receptor (TLR) signaling. Periodontitis is one of the most common inflammatory diseases in humans that results in the destruction of periodontal tissues and alveolar bone, which ultimately leads to teeth loss. It is estimated that majority of adults over the age of 30 suffer from periodontal bone loss. Growing evidence suggests that chronic periodontal inflammation is an important risk factor for several pathological disorders including cardiovascular disease, diabetes, atherosclerosis and arthritis. Thus, periodontitis is a significant public health concern particularly to aged people. Consequently, there is still an urgent need for developing better treatments and preventative strategies that can dramatically reduce the inflammation, bone loss, and systemic ramifications of periodontitis. The current lack of highly effective therapies may be largely due to incomplete knowledge of the mechanism of periodontitis pathogenesis. Surprisingly, although TLRs are critical for host defense and inflammatory diseases, the role of TLRs in the pathogenesis of periodontitis remains largely unknown. We found that adeno-associated virus (AAV) Ctsk shRNAi (AAV-shRNA-Ctsk) mediated silencing prevents both bone loss and inflammation in a mouse model of endodontic disease. Moreover, it was reported that Ctsk is required for TLR9 signaling in a rat model of rheumatoid arthritis. Notably, our preliminary data showed that Cathepsin K gene knockout and AAV Ctsk shRNA (AAV-shRNA-Ctsk) mediated silencing dramatically prevents both bone loss and inflammation in a mouse model of periodontitis. Collectively, these studies strongly indicate that Ctsk is a major osteoimmune gene that can be targeted to control both inflammation and bone loss, and that Ctsk may be a key regulator of TLRs signaling. Based on our studies and those of others, we hypothesize that targeting Ctsk inhibits inflammation and bone loss caused by bacteria infection in periodontitis through attenuation of TLRs signaling. Three specific aims are proposed to test our hypothesis. We will define the functional role of Ctsk in the TLRs signaling-mediated immune response and bone resorption induced by periodontitis through comparative analysis of Ctsk-/-, Ctsk+/-, and Ctsk+/+ mice infected with periodontal pathogens in Aim 1. We will determine the therapeutic potential of AAV-shRNA-Ctsk as a means to reduce the progression and severity of periodontitis in vivo by attenuating TLRs signaling in Aim 2. We will characterize the mechanism by which Ctsk mediates TLRs signaling induced by pathogens and inflammatory mediators in a mouse periodontitis model using dendritic cells, fibroblasts and macrophages in Aim 3. A multidisciplinary research team (i.e. a molecular geneticist, dentist scientists, animal model experts and an immunologist) has been established to achieve the research goal. This study will not only improve our understanding mechanism of basic knowledge of the pathogenesis of periodontitis, but it will facilitate the design of novel therapeutic approaches fo this disease.